The Hox genes, a highly conserved family of transcription factors, are expressed within specific anterior-posterior domains of the central nervous system in all animal species examined thus far. Nine Hox genes have been cloned and partially characterized in the medicinal leech Hirudo medicinalis. These genes are expressed in different but partially overlapping sets of neurons, where they are likely to control the establishment of neuronal identities during development. The main goal of this project is to test this hypothesis in the leech, which is a very suitable system for the study of the individual features of neurons at the single-cell level. Using antibodies raised against the proteins produced by these genes, their expression patters will be studied with detail. The extent and nature of the overlap in the expression domains of Hox genes will be analyzed within each ganglion. Using markers of neuronal types that are available in the leech system and dye injections, neurons that express the same Hox gene will be studied in order to determine whether they share particular characteristics, such as neurotransmitter type, expression of marker molecules, or axonal morphology. The functions of these genes will be further investigated by infecting antisense oligonucleotides into the body cavity of embryos, to block the expression of individual Hox genes. The effects of blocking Hox gene expression during the development of the central nervous system will be studied in identified neurons. Those experiments provide an opportunity to study the relationships between Hox gene expression and the acquisition of specific properties by individual neurons, an opportunity that is largely unavailable in other systems.